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1.
Bioorg Chem ; 105: 104429, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33161256

RESUMO

Human serum albumin (HSA) as the most abundant protein in human blood plasma, can be a good indicator for evaluating severity of some diseases in the clinic. HSA can be find in two forms: reduced albumin (human mercaptalbumin (HMA)) and oxidized albumin (human non-mercaptalbumin (HNA)). The rate of oxidized albumin to total albumin can be enhanced in multiple diseases. Increase in HNA level have been demonstrated in liver, diabetes plus fatigue and coronary artery diseases. In liver patients, this enhancement can reach to 50-200 percent which can then lead to bacterial/viral infections and eventually death in severe conditions. Due to the induction of cytokine storm, we can say that the level of HNA in serum of coronavirus disease 2019 (COVID-19) patients may be a positive predictor of mortality, especially in patients with underlying diseases such as cardiovascular disease (CVD), diabetes, aging and other inflammatory diseases. We suggest that checking oxidized albumin in COVID-19 patients may provide new therapeutic and diagnostic opportunities to better combat COVID-19.


Assuntos
/diagnóstico , Albumina Sérica Humana/análise , /terapia , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Fígado/metabolismo , Oxirredução , Espécies Reativas de Oxigênio/química , Espécies Reativas de Oxigênio/metabolismo , Albumina Sérica/análise , Albumina Sérica/química , Albumina Sérica Humana/química
2.
PLoS One ; 15(11): e0239938, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33166313

RESUMO

Mycobacterium bovis, the causative agent of bovine tuberculosis, is a pathogen that impacts both animal and human health. Consequently, there is a need to improve understanding of disease dynamics, identification of infected animals, and characterization of the basis of immune protection. This study assessed the transcriptional changes occurring in cattle during the early weeks following a M. bovis infection. RNA-seq analysis of whole blood-cell transcriptomes revealed two distinct transcriptional clusters of infected cattle at both 4- and 10-weeks post-infection that correlated with disease severity. Cattle exhibiting more severe disease were transcriptionally divergent from uninfected animals. At 4-weeks post-infection, 25 genes had commonly increased expression in infected cattle compared to uninfected cattle regardless of disease severity. Ten weeks post-infection, differential gene expression was only observed when severely-affected cattle were compared to uninfected cattle. This indicates a transcriptional divergence based on clinical status following infection. In cattle with more severe disease, biological processes and cell type enrichment analyses revealed overrepresentation of innate immune-related processes and cell types in infected animals. Collectively, our findings demonstrate two distinct transcriptional profiles occur in cattle following M. bovis infection, which correlate to clinical status.


Assuntos
Imunidade Inata/genética , Leucócitos Mononucleares/metabolismo , Mycobacterium bovis/imunologia , Transcriptoma/genética , Tuberculose Bovina/patologia , Animais , Bovinos , Expressão Gênica/genética , Perfilação da Expressão Gênica/veterinária , Leucócitos Mononucleares/citologia , Mycobacterium bovis/genética , Análise de Sequência com Séries de Oligonucleotídeos , Índice de Gravidade de Doença , Tuberculose Bovina/imunologia
3.
Cell Death Dis ; 11(11): 957, 2020 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-33159040

RESUMO

A global effort is currently undertaken to restrain the COVID-19 pandemic. Host immunity has come out as a determinant for COVID-19 clinical outcomes, and several studies investigated the immune profiling of SARS-CoV-2 infected people to properly direct the clinical management of the disease. Thus, lymphopenia, T-cell exhaustion, and the increased levels of inflammatory mediators have been described in COVID-19 patients, in particular in severe cases1. Age represents a key factor in COVID-19 morbidity and mortality2. Understanding age-associated immune signatures of patients are therefore important to identify preventive and therapeutic strategies. In this study, we investigated the immune profile of COVID-19 hospitalized patients identifying a distinctive age-dependent immune signature associated with disease severity. Indeed, defined circulating factors - CXCL8, IL-10, IL-15, IL-27, and TNF-α - positively correlate with older age, longer hospitalization, and a more severe form of the disease and may thus represent the leading signature in critical COVID-19 patients.


Assuntos
Infecções por Coronavirus/patologia , Citocinas/metabolismo , Pneumonia Viral/patologia , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Betacoronavirus/imunologia , Betacoronavirus/isolamento & purificação , Análise por Conglomerados , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Feminino , Humanos , Imunoglobulina G/sangue , Interleucina-10/metabolismo , Interleucina-8/metabolismo , Tempo de Internação , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Índice de Gravidade de Doença , Fator de Necrose Tumoral alfa/metabolismo
4.
PLoS One ; 15(10): e0240335, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33057374

RESUMO

BACKGROUND: Increased acute rejection risk in rescue protocols with Belatacept may limit its use particularly in medically complex patients where preexisting increased risk of rejection couples with CNI toxicity. METHODS: Retrospective analysis was performed in 19 KTs shifted to a Belatacept-based immunosuppression with low-dose Tacrolimus (2-3 ng/mL) after evidence of allograft disfunction, including patients with primary non-function (PNF), chronic-active antibody-mediated rejection (cAMR), history of previous KTs and/or other concomitant transplants (liver, pancreas). Evaluation of CD28+ CD4+ effector memory T cell (TEM) before conversion was performed in 10/19. RESULTS: Kidney function significantly improved (median eGFR 16.5 ml/min/1.73m2 before vs 25 ml/min after; p = 0.001) at a median time after conversion of 12.5 months (9.1-17.8). Overall graft and patient survival were 89.5% and 100% respectively. Definitive weaning from dialysis in 5/5 KTs with PNF was observed, whereas 7/8 patients lost their graft within first year in a control group. eGFR significantly ameliorated in re-trasplants (p = 0.001) and stabilized in KTs with other organ transplants or cAMR. No acute rejection episodes occurred, despite the significant risk suggested by high frequency of CD28+ CD4+ TEM in most patients. Opportunistic infections were limited and most common in early vs late-converted. CONCLUSIONS: Rescue association of Belatacept with low-dose Tacrolimus in medically complex KTs is a feasible option that allows prevention of acute rejection and amelioration of graft function.


Assuntos
Abatacepte/uso terapêutico , Rejeição de Enxerto/prevenção & controle , Imunossupressores/uso terapêutico , Tacrolimo/uso terapêutico , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Feminino , Taxa de Filtração Glomerular , Rejeição de Enxerto/etiologia , Sobrevivência de Enxerto , Humanos , Transplante de Rim/efeitos adversos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Fenótipo , Estudos Retrospectivos , Transplante Homólogo
5.
J Med Chem ; 63(21): 12921-12928, 2020 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-33126797

RESUMO

Therapeutic options to treat multidrug resistant bacteria, especially when present in biofilms, are limited due to their high levels of antibiotic resistance. Here, we report the anti-biofilm and immunomodulatory activities of the host defense peptide (HDP)-mimicking ß-peptide polymer (20:80 Bu:DM) and investigated its activity in vivo. The polymer outperformed antibiotics in the removal and reduction of the viability of established biofilms, achieving a maximum activity of around 80% reduction in viability. Interestingly the polymer also exhibited HDP-like immunomodulation in inducing chemokines and anti-inflammatory cytokines and suppressing lipopolysaccharide-induced proinflammatory cytokines. When tested in a murine, high-density skin infection model using P. aeruginosa LESB58, the polymer was effective in diminishing abscess size and reducing bacterial load. This study demonstrates the dual functionality of HDP-mimicking ß-peptide polymers in inhibiting biofilms and modulating innate immunity, as well as reducing tissue dermonecrosis.


Assuntos
Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Biofilmes/efeitos dos fármacos , Polímeros/química , Animais , Antibacterianos/farmacologia , Anti-Infecciosos/química , Anti-Infecciosos/uso terapêutico , Sobrevivência Celular/efeitos dos fármacos , Citocinas/farmacologia , Modelos Animais de Doenças , Humanos , Imunidade Inata/efeitos dos fármacos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Staphylococcus aureus Resistente à Meticilina , Testes de Sensibilidade Microbiana , Peptidomiméticos , Polímeros/farmacologia , Polímeros/uso terapêutico , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , Dermatopatias/tratamento farmacológico , Dermatopatias/microbiologia , Dermatopatias/patologia
6.
Sci Rep ; 10(1): 14094, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32839476

RESUMO

The aim of this study was to analyze the metabolic and molecular profile according to physical fitness status (Low or High VO2max) and its impacts on peripheral and cellular inflammatory responses in healthy men. First (Phase I), inflammatory profile (TNF-α, IL-6, IL-10) was analyzed at baseline and post-acute exercise sessions performed at low (< 60% VO2max) and high (> 90% VO2max) intensities considering the individual endotoxin concentrations. Next (Phase II), monocyte cell cultures were treated with LPS alone or associated with Rosiglitazone (PPAR-γ agonist drug) to analyze cytokine production and gene expression. Monocyte subsets were also evaluated by flow cytometry. A positive relationship was observed between LPS concentrations and oxygen uptake (VO2max) (r = 0.368; p = 0.007); however, in the post-exercise an inverse correlation was found between LPS variation (Δ%) and VO2max (r = -0.385; p = 0.004). With the low-intensity exercise session, there was inverse correlation between LPS and IL-6 concentrations post-exercise (r = -0.505; p = 0.046) and a positive correlation with IL-10 in the recovery (1 h post) (r = 0.567; p = 0.011), whereas with the high-intensity exercise an inverse correlation was observed with IL-6 at pre-exercise (r = -0.621; p = 0.013) and recovery (r = -0.574; p = 0.016). When monocyte cells were treated with LPS, High VO2max individuals showed higher PPAR-γ gene expression whereas Low VO2max individuals displayed higher IL-10 production. Additionally, higher TLR-4, IKK1, and PGC-1α gene expression were observed in the High VO2max group than Low VO2max individuals. In conclusion, even with elevated endotoxemia, individuals with High VO2max exhibited higher IL-6 concentration in peripheral blood post-acute aerobic exercise and lower IL-10 concentration during recovery (1 h post-exercise). The anti-inflammatory effects linked with exercise training and physical fitness status may be explained by a greater gene expression of IKK1, TLR-4, and PGC-1α, displaying an extremely efficient cellular framework for the PPAR-γ responses.


Assuntos
Exercício Físico/fisiologia , Lipopolissacarídeos/metabolismo , Consumo de Oxigênio/fisiologia , PPAR gama/sangue , PPAR gama/metabolismo , Aptidão Física/fisiologia , Adulto , Células Cultivadas , Endotoxemia/sangue , Humanos , Hipoglicemiantes/farmacologia , Interleucina-10/sangue , Interleucina-6/sangue , Leucócitos Mononucleares/citologia , Lipopolissacarídeos/sangue , Masculino , PPAR gama/agonistas , Rosiglitazona/farmacologia , Fator de Necrose Tumoral alfa/sangue
7.
J Med Chem ; 63(17): 9020-9044, 2020 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-32787145

RESUMO

The bromodomain and extraterminal domain (BET) family of epigenetic regulators comprises four proteins (BRD2, BRD3, BRD4, BRDT), each containing tandem bromodomains. To date, small molecule inhibitors of these proteins typically bind all eight bromodomains of the family with similar affinity, resulting in a diverse range of biological effects. To enable further understanding of the broad phenotype characteristic of pan-BET inhibition, the development of inhibitors selective for individual, or sets of, bromodomains within the family is required. In this regard, we report the discovery of a potent probe molecule possessing up to 150-fold selectivity for the N-terminal bromodomains (BD1s) over the C-terminal bromodomains (BD2s) of the BETs. Guided by structural information, a specific amino acid difference between BD1 and BD2 domains was targeted for selective interaction with chemical functionality appended to the previously developed I-BET151 scaffold. Data presented herein demonstrate that selective inhibition of BD1 domains is sufficient to drive anti-inflammatory and antiproliferative effects.


Assuntos
Anti-Inflamatórios/química , Proteínas de Ciclo Celular/antagonistas & inibidores , Desenho de Fármacos , Fatores de Transcrição/antagonistas & inibidores , Animais , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/farmacologia , Sítios de Ligação , Proteínas de Ciclo Celular/classificação , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Citocinas/metabolismo , Meia-Vida , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Masculino , Camundongos , Simulação de Dinâmica Molecular , Filogenia , Domínios Proteicos , Quinolonas/química , Quinolonas/metabolismo , Quinolonas/farmacologia , Fatores de Transcrição/classificação , Fatores de Transcrição/metabolismo
8.
EMBO Mol Med ; 12(10): e13038, 2020 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-32816392

RESUMO

Early in the COVID-19 pandemic, type 2 diabetes (T2D) was marked as a risk factor for severe disease and mortality. Inflammation is central to the aetiology of both conditions where variations in immune responses can mitigate or aggravate disease course. Identifying at-risk groups based on immunoinflammatory signatures is valuable in directing personalised care and developing potential targets for precision therapy. This observational study characterised immunophenotypic variation associated with COVID-19 severity in T2D. Broad-spectrum immunophenotyping quantified 15 leucocyte populations in peripheral circulation from a cohort of 45 hospitalised COVID-19 patients with and without T2D. Lymphocytopenia and specific loss of cytotoxic CD8+ lymphocytes were associated with severe COVID-19 and requirement for intensive care in both non-diabetic and T2D patients. A morphological anomaly of increased monocyte size and monocytopenia restricted to classical CD14Hi CD16- monocytes was specifically associated with severe COVID-19 in patients with T2D requiring intensive care. Increased expression of inflammatory markers reminiscent of the type 1 interferon pathway (IL6, IL8, CCL2, INFB1) underlaid the immunophenotype associated with T2D. These immunophenotypic and hyperinflammatory changes may contribute to increased voracity of COVID-19 in T2D. These findings allow precise identification of T2D patients with severe COVID-19 as well as provide evidence that the type 1 interferon pathway may be an actionable therapeutic target for future studies.


Assuntos
/patologia , Diabetes Mellitus Tipo 2/patologia , Monócitos/fisiologia , Idoso , /virologia , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Diabetes Mellitus Tipo 2/complicações , Feminino , Humanos , Imunofenotipagem , Inflamação/etiologia , Interleucina-6/genética , Interleucina-6/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Receptores de Lipopolissacarídeos/metabolismo , Linfopenia/diagnóstico , Masculino , Pessoa de Meia-Idade , Monócitos/citologia , Monócitos/patologia , Fatores de Risco , Índice de Gravidade de Doença
9.
Chem Biol Interact ; 329: 109223, 2020 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-32781033

RESUMO

Thromboembolism is a major cause of morbidity and mortality worldwide. Most therapeutic drugs for treating thrombosis can cause hemorrhage and have short half-lives within human blood circulation resulting in a need to discover and develop novel anticoagulants/antithrombotics. EuRP-61 has been isolated from a plant latex (Euphorbia resinifera) and characterized as a serine protease. In this study, EuRP-61 was able to hydrolyze all chains of human fibrin clots. The enzyme may have long term stability in blood circulation as its fibrinogenolytic activity was not affected by human blood circulating inhibitors such as α2-macroglobulin and antithrombin III. The enzyme may affect the extrinsic, intrinsic or common pathways of the human blood coagulation cascade as evidenced by its prolonged of both prothrombin (PT) and activated partial thromboplastin (APTT) time. Moreover, the enzyme inhibited platelet aggregation via the ADP-receptor pathway. EuRP-61 was not toxic to human red blood cells in the 4 common blood groups (A, B, O and AB) (all Rh+) or human peripheral blood mononuclear cells (hPBMCs). The enzyme may protect human peripheral blood cells from aggregation without destroying them. This study provides evidence that EuRP-61 may have potential as an agent for the treatment of thrombosis.


Assuntos
Anticoagulantes/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Euphorbia/enzimologia , Fibrinolíticos/farmacologia , Peptídeo Hidrolases/farmacologia , Proteínas de Plantas/farmacologia , Anticoagulantes/isolamento & purificação , Antitrombina III/antagonistas & inibidores , Antitrombina III/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Fibrinolíticos/isolamento & purificação , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Peptídeo Hidrolases/isolamento & purificação , Proteínas de Plantas/isolamento & purificação , Agregação Plaquetária/efeitos dos fármacos , alfa 2-Macroglobulinas Associadas à Gravidez/antagonistas & inibidores , alfa 2-Macroglobulinas Associadas à Gravidez/metabolismo
10.
BMC Infect Dis ; 20(1): 590, 2020 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-32778058

RESUMO

BACKGROUND: Antiviral therapy is recommended for patients with immune-active chronic hepatitis B (CHB) to decrease the risk of liver-related complications. However, the outcomes of the pegylated IFN-α (PEG-IFN-α) therapy vary among CHB patients. We aimed to identify factors that can influence the outcomes in CHB patients who received antiviral PEG-IFN-α monotherapy. METHODS: Thirty-two CHB patients who received PEG-IFN-α monotherapy were enrolled in this study. All of the patients underwent two liver biopsies at baseline and 6 months after the initiation of the therapy. CD8+ T cells, CD4+ T cells, CD68+ mononuclear cells, and PD-1 levels in the 64 liver biopsy specimens were examined via immunofluorescence. RESULTS: The overall median frequency of CD8+ T cells in the liver tissues of 32 CHB patients significantly decreased at 6 months after the therapy initiation (p < 0.01). In the FIER (fibrosis and inflammation response with HBeAg seroconversion) group, CD8+PD-1+ T cells significantly decreased at 6 months (p < 0.05), while CD8+PD-1- T cells had no significant difference. On the contrary, in the FIENR (no fibrosis and inflammation response and HBeAg seroconversion) group, CD8+PD-1- T cells significantly decreased after 6 months of PEG-IFN-α treatment (p < 0.05), while CD8+PD-1+ T cells had no significant difference. In addition, the levels of CD68+ mononuclear cells in the FIER group showed an overall increasing trend after treatment (p < 0.05). CONCLUSIONS: The changes in the levels of CD8+PD-1+ T cells and CD68+ mononuclear cells may be related to the response to PEG-IFN-α therapy.


Assuntos
Antivirais/uso terapêutico , Linfócitos T CD8-Positivos/metabolismo , Hepatite B Crônica/tratamento farmacológico , Interferon-alfa/uso terapêutico , Fígado/patologia , Polietilenoglicóis/uso terapêutico , Adulto , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Antígenos E da Hepatite B/sangue , Antígenos E da Hepatite B/imunologia , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Fígado/metabolismo , Masculino , Receptor de Morte Celular Programada 1/metabolismo , Proteínas Recombinantes/uso terapêutico , Adulto Jovem
11.
Arch Med Res ; 51(7): 645-653, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32611485

RESUMO

BACKGROUND: The SARS-CoV-2 is the etiological agent causing COVID-19 which has infected more than 2 million people with more than 200000 deaths since its emergence in December 2019. In the majority of cases patients are either asymptomatic or show mild to moderate symptoms and signs of a common cold. A subset of patients, however, develop a severe atypical pneumonia, with the characteristic ground-glass appearance on chest x-ray and computerized tomography, which evolves into an acute respiratory distress syndrome, that requires mechanical ventilation and eventually results in multiple organ failure and death. The Molecular pathogenesis of COVID-19 is still unknown. AIM OF THE STUDY: In the present work we performed a stringent metanalysis from the publicly available RNAseq data from bronchoalveolar cells and peripheral blood mononuclear cells to elucidate molecular alterations and cellular deconvolution to identify immune cell profiles. RESULTS: Alterations in genes involved in hyaluronan, glycosaminoglycan and mucopolysaccharides metabolism were over-represented in bronchoalveolar cells infected by SARS-CoV-2, as well as potential lung infiltration with neutrophils, T CD4+ cell and macrophages. The blood mononuclear cells presented a proliferative state. Dramatic reduction of NK and T lymphocytes, whereas an exacerbated increase in monocytes. CONCLUSIONS: In summary our results revealed molecular pathogenesis of the SARS-CoV-2 infection to bronchoalveolar cells inducing the hyaluronan and glycosaminoglycan metabolism that could shape partially the components of the ground-glass opacities observed in CT. And the potential immune response profile in COVID-19.


Assuntos
Glicosaminoglicanos , Líquido da Lavagem Broncoalveolar/citologia , /genética , /patologia , Glicosaminoglicanos/genética , Glicosaminoglicanos/metabolismo , Humanos , Ácido Hialurônico/genética , Ácido Hialurônico/metabolismo , Leucócitos Mononucleares/citologia , Pulmão/diagnóstico por imagem , Pulmão/patologia
12.
PLoS One ; 15(7): e0235518, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32614928

RESUMO

Interruption of the programmed death 1 (PD-1) / programmed death ligand 1 (PD-L1) pathway is an established and effective therapeutic strategy in human oncology and holds promise for veterinary oncology. We report the generation and characterization of monoclonal antibodies specific for canine PD-1 and PD-L1. Antibodies were initially assessed for their capacity to block the binding of recombinant canine PD-1 to recombinant canine PD-L1 and then ranked based on efficiency of binding as judged by flow cytometry. Selected antibodies were capable of detecting PD-1 and PD-L1 on canine tissues by flow cytometry and Western blot. Anti-PD-L1 worked for immunocytochemistry and anti-PD-1 worked for immunohistochemistry on formalin-fixed paraffin embedded canine tissues, suggesting the usage of this antibody with archived tissues. Additionally, anti-PD-L1 (JC071) revealed significantly increased PD-L1 expression on canine monocytes after stimulation with peptidoglycan or lipopolysaccharide. Together, these antibodies display specificity for the natural canine ligand using a variety of potential diagnostic applications. Importantly, multiple PD-L1-specific antibodies amplified IFN-γ production in a canine peripheral blood mononuclear cells (PBMC) concanavlin A (Con A) stimulation assay, demonstrating functional activity.


Assuntos
Anticorpos Monoclonais/imunologia , Antígeno B7-H1/imunologia , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T/imunologia , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Cães , Interferon gama/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Peptidoglicano/farmacologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/metabolismo
13.
Sci Rep ; 10(1): 12200, 2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32699258

RESUMO

In this study, we compared the effect of tricarbonyldichlororuthenium (II) dimer (CORM-2) and its CO-depleted molecule (iCORM-2) on human peripheral blood mononuclear cells (PBMCs) and human promyelocytic leukemia HL-60 cells. We determined cell viability, DNA damage and DNA repair kinetics. We also studied the effect of both compounds on DNA oxidative damage, free radical level and HO-1 gene expression. We showed that at low concentrations both CORM-2 and iCORM-2 stimulate PBMCs viability. After 24-h incubation, CORM-2 and iCORM-2, at the concentration of 100 µM, reduce the viability of both PBMCs and HL-60 cells. We also demonstrated that CORM-2 and iCORM-2, in the 0.01-100 µM concentration range, cause DNA damage such as strand breaks and alkaline labile sites. DNA damage was repaired efficiently only in HL-60 cells. CORM-2 significantly reduces oxidative stress induced by 1 mM H2O2 in normal and cancer cells. On the contrary, iCORM-2 in HL-60 cells increases the level of free radicals in the presence of 1 and 5 mM H2O2. We also revealed that both CORM-2 and iCORM-2 induce HO-1 gene expression. However, CORM-2 induces this gene to a greater extent than iCORM-2, especially in HL-60 cells at 100 µM. Finally, we showed that CORM-2 and iCORM-2 reduce H2O2-induced DNA oxidative damage. Furthermore, CORM-2 proved to be a compound with stronger antioxidant properties than iCORM-2. Our results suggest that both active CORM-2 and inactive iCORM-2 exert biological effects such as cyto- and genotoxicity, antioxidant properties and the ability to induce the HO-1 gene. The released CO as well as iCORM-2 can be responsible for these effects.


Assuntos
Antioxidantes/química , Monóxido de Carbono/química , Dano ao DNA/efeitos dos fármacos , Compostos Organometálicos/farmacologia , Monóxido de Carbono/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Radicais Livres/metabolismo , Células HL-60 , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Humanos , Peróxido de Hidrogênio/química , Peróxido de Hidrogênio/farmacologia , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
14.
Sci Rep ; 10(1): 12098, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694530

RESUMO

Chloroquine (CQ) and hydroxychloroquine, are promising anti-inflammatory drugs for the treatment of Diabetes mellitus (DM) to prevent associated complications. Therefore, this study evaluated the anti-inflammatory effects of CQ-free and CQ-incorporated polylactic acid nanoparticles (NPs) in the peripheral blood mononuclear cells (PBMCs) of patients with type 1 Diabetes mellitus (T1DM). In total, 25 normoglycemic individuals and 25 patients with T1DM aged 10-16 years were selected and glycemic controls evaluated. After cell viability assessed by MTT assay, T1DM PBMCs were subjected to a CQ concentration of 10 µM in three different conditions: not treated (NT), treated with CQ, and treated with CQ NPs. The cells were incubated for 48 h, and the mRNA expressions of cytokines IL1B, IFNG, TNFA, IL12, and IL10 were determined by relative quantification through real-time PCR at 24 h intervals. IL1B expression decreased in CQ and CQ NP-treated cells after 48 h (p < 0.001) and 24 h (p < 0.05) of treatment, respectively. IFNG and IL12 expressions significantly decreased (p < 0.001) in cells treated with CQ and CQ NPs at 24 and 48 h compared to NT. TNFA and IL10 expressions significantly decreased after 48 h (p < 0.001) and 24 h (p < 0.002), respectively, by both CQ and CQ NPs treatment. Despite being a preliminary in vitro study, CQ has anti-inflammatory activity in the primary cells of T1DM patients and could represent an alternative and adjuvant anti-inflammatory therapy to prevent diabetes complications.


Assuntos
Anti-Inflamatórios/farmacologia , Cloroquina/farmacologia , Citocinas/genética , Diabetes Mellitus Tipo 1/genética , Leucócitos Mononucleares/citologia , Poliésteres/química , Adolescente , Anti-Inflamatórios/química , Estudos de Casos e Controles , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Quimioterapia Adjuvante , Criança , Cloroquina/química , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/imunologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Masculino , Nanopartículas
15.
Sci Rep ; 10(1): 12130, 2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32699287

RESUMO

The evidence for the beneficial effects of drinking hydrogen-water (HW) is rare. We aimed to investigate the effects of HW consumption on oxidative stress and immune functions in healthy adults using systemic approaches of biochemical, cellular, and molecular nutrition. In a randomized, double-blind, placebo-controlled study, healthy adults (20-59 y) consumed either 1.5 L/d of HW (n = 20) or plain water (PW, n = 18) for 4 weeks. The changes from baseline to the 4th week in serum biological antioxidant potential (BAP), derivatives of reactive oxygen, and 8-Oxo-2'-deoxyguanosine did not differ between groups; however, in those aged ≥ 30 y, BAP increased greater in the HW group than the PW group. Apoptosis of peripheral blood mononuclear cells (PBMCs) was significantly less in the HW group. Flow cytometry analysis of CD4+, CD8+, CD20+, CD14+ and CD11b+ cells showed that the frequency of CD14+ cells decreased in the HW group. RNA-sequencing analysis of PBMCs demonstrated that the transcriptomes of the HW group were clearly distinguished from those of the PW group. Most notably, transcriptional networks of inflammatory responses and NF-κB signaling were significantly down-regulated in the HW group. These finding suggest HW increases antioxidant capacity thereby reducing inflammatory responses in healthy adults.


Assuntos
Apoptose , Hidrogênio/química , Leucócitos Mononucleares/metabolismo , Água/administração & dosagem , Adulto , Antioxidantes/análise , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Análise por Conglomerados , Método Duplo-Cego , Regulação para Baixo/efeitos dos fármacos , Feminino , Voluntários Saudáveis , Humanos , Hidrogênio/administração & dosagem , Hidrogênio/farmacologia , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Receptores de Lipopolissacarídeos/metabolismo , Masculino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Efeito Placebo , Transcriptoma , Água/química , Adulto Jovem
16.
J Vis Exp ; (160)2020 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-32597837

RESUMO

Studying isolated cells from mucosa-associated lymphoid tissues (MALT) allows understanding of immune cells response in pathologies involving mucosal immunity, because they can model host-pathogen interactions in the tissue. While isolated cells derived from tissues were the first cell culture model, their use has been neglected because tissue can be hard to obtain. In the present protocol, we explain how to easily process and culture tonsillar mononuclear cells (TMCs) from healthy human tonsils to study innate immune responses upon activation, mimicking viral infection in mucosal tissues. Isolation of TMCs from the tonsils is quick, because the tonsils barely have any epithelium and yield up to billions of all major immune cell types. This method allows detection of cytokine production using several techniques, including immunoassays, qPCR, microscopy, flow cytometry, etc., similar to the use of peripheral mononuclear cells (PBMCs) from blood. Furthermore, TMCs show a higher sensitivity to drug testing than PBMCs, which needs to be considered for future toxicity assays. Thus, ex vivo TMCs cultures are an easy and accessible mucosal model.


Assuntos
Separação Celular/métodos , Imunidade Inata/imunologia , Imunidade nas Mucosas/imunologia , Leucócitos Mononucleares/citologia , Tecido Linfoide/imunologia , Tonsila Palatina/citologia , Citometria de Fluxo , Humanos , Leucócitos Mononucleares/imunologia , Tecido Linfoide/citologia , Tonsila Palatina/imunologia
17.
PLoS One ; 15(6): e0235422, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584899

RESUMO

Alternatives to antibiotics for prevention of respiratory tract infections in cattle are urgently needed given the increasing public and regulatory pressure to reduce overall antibiotic usage. Activation of local innate immune defenses in the upper respiratory tract is one strategy to induce non-specific protection against infection with the diverse array of viral and bacterial pathogens associated with bovine respiratory disease complex (BRDC), while avoiding the use of antibiotics. Our prior studies in rodent models demonstrated that intranasal administration of liposome-TLR complexes (LTC) as a non-specific immune stimulant generated high levels of protection against lethal bacterial and viral pathogens. Therefore, we conducted studies to assess LTC induction of local immune responses and protective immunity to BRDC in cattle. In vitro, LTC were shown to activate peripheral blood mononuclear cells in cattle, which was associated with secretion of INFγ and IL-6. Macrophage activation with LTC triggered intracellular killing of Mannheimia hemolytica and several other bacterial pathogens. In studies in cattle, intranasal administration of LTC demonstrated dose-dependent activation of local innate immune responses in the nasopharynx, including recruitment of monocytes and prolonged upregulation (at least 2 weeks) of innate immune cytokine gene expression by nasopharyngeal mucosal cells. In a BRDC challenge study, intranasal administration of LTC prior to pathogen exposure resulted in significant reduction in both clinical signs of infection and disease-associated euthanasia rates. These findings indicate that intranasal administration of a non-specific innate immune stimulant can be an effective method of rapidly generating generalized protection from mixed viral and bacterial respiratory tract infections in cattle.


Assuntos
Complexo Respiratório Bovino/patologia , Imunidade Inata/efeitos dos fármacos , Fármacos do Sistema Respiratório/farmacologia , Administração Intranasal , Animais , Complexo Respiratório Bovino/tratamento farmacológico , Complexo Respiratório Bovino/mortalidade , Bovinos , Antígenos de Histocompatibilidade Classe II/metabolismo , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Lipossomos/química , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Mannheimia haemolytica/isolamento & purificação , Mannheimia haemolytica/patogenicidade , Nasofaringe/metabolismo , Nasofaringe/microbiologia , Óxido Nítrico/metabolismo , Fagocitose , Fármacos do Sistema Respiratório/uso terapêutico , Taxa de Sobrevida , Receptor 3 Toll-Like/agonistas , Receptor Toll-Like 9/agonistas , Regulação para Cima/efeitos dos fármacos
18.
PLoS One ; 15(6): e0233980, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32511247

RESUMO

Multiple sclerosis (MS) is an inflammatory and demyelinating disease of the central nervous system (CNS) that results in variable severities of neurodegeneration. The understanding of MS has been limited by the inaccessibility of the affected cells and the lengthy timeframe of disease development. However, recent advances in stem cell technology have facilitated the bypassing of some of these challenges. Towards gaining a greater understanding of the innate potential of stem cells from people with varying degrees of disability, we generated induced pluripotent stem cells (iPSCs) from peripheral blood mononuclear cells derived from stable and progressive MS patients, and then further differentiated them into oligodendrocyte (OL) lineage cells. We analyzed differentiation under both homeostatic and inflammatory conditions via sustained exposure to low-dose interferon gamma (IFNγ), a prominent cytokine in MS. We found that all iPSC lines differentiated into mature myelinating OLs, but chronic exposure to IFNγ dramatically inhibited differentiation in both MS groups, particularly if exposure was initiated during the pre-progenitor stage. Low-dose IFNγ was not toxic but led to an early upregulation of interferon response genes in OPCs followed by an apparent redirection in lineage commitment from OL to a neuron-like phenotype in a significant portion of the treated cells. Our results reveal that a chronic low-grade inflammatory environment may have profound effects on the efficacy of regenerative therapies.


Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Esclerose Múltipla Crônica Progressiva/patologia , Oligodendroglia/citologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Homeostase , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/fisiologia , Inflamação/patologia , Interferon gama/farmacologia , Leucócitos Mononucleares/citologia , Regeneração
19.
BJU Int ; 126(3): 379-387, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32437049

RESUMO

OBJECTIVES: To assess the presence of self-reactive immune responses to seminal and prostate antigens (PAg), biomarkers of inflammation of the male genital tract, and semen quality parameters in patients with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS). PATIENTS, SUBJECTS AND METHODS: Peripheral blood and semen samples were collected from patients with CP/CPPS and age-matched healthy control volunteers. We analysed the lymphoproliferative responses of peripheral blood mononuclear cells (PBMC) to different seminal plasma (SP)-derived and purified PAg, serum autoantibodies specific to PAg, leucocyte subpopulations, and inflammatory cytokines in semen, sperm apoptosis/necrosis, and semen quality parameters. RESULTS: Significantly greater PBMC proliferative responses specific to PAg, with elevated secretion of interferon (IFN)γ and interleukin (IL)-17, were detected in the patients with CP/CPPS vs the controls. Moreover, the patients with CP/CPPS had significantly greater serum immunoglobulin G immune reactivity to SP proteins, such as prostate-specific antigen and prostatic acid phosphatase, than the controls. Inflammation of the male genital tract was exemplified by high levels of IFNγ, IL-17, IL-1ß and IL-8, as well as higher counts of leukocytes, mainly CD4 T lymphocytes and macrophages, in the semen. In addition, this local inflammation was associated with an overall diminished semen quality, i.e., reduced sperm concentration, motility and viability; and higher levels of sperm apoptosis/necrosis in patients with CP/CPPS vs controls. CONCLUSION: Patients with CP/CPPS show T helper type 1 (Th1) and Th17 immune responses specific to PAg associated with chronic inflammation of the male genital tract and reduced semen quality. These immune responses may underlie the induction and development of chronic pelvic pain and inflammation of the male genital tract, which in turn could alter normal prostate functioning and impair semen quality.


Assuntos
Autoantígenos/imunologia , Próstata/imunologia , Prostatite/imunologia , Prostatite/fisiopatologia , Análise do Sêmen , Sêmen/imunologia , Células Th1/imunologia , Células Th17/imunologia , Adulto , Proliferação de Células , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Prostatite/sangue
20.
BMC Genomics ; 21(1): 350, 2020 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393182

RESUMO

BACKGROUND: The thymus is a highly specialized organ of the immune system where T cell precursors develop and differentiate into self-tolerant CD4+ or CD8+ T cells. No studies to date have investigated how the human transcriptome profiles differ, between T cells still residing in the thymus and T cells in the periphery. RESULTS: We have performed high-throughput RNA sequencing to characterize the transcriptomes of primary single positive (SP) CD4+ and CD8+ T cells from infant thymic tissue, as well as primary CD4+ and CD8+ T cells from infant and adult peripheral blood, to enable the comparisons across tissues and ages. In addition, we have assessed the expression of candidate genes related to autoimmune diseases in thymic CD4+ and CD8+ T cells. The thymic T cells showed the largest number of uniquely expressed genes, suggesting a more diverse transcription in thymic T cells. Comparing T cells of thymic and blood origin, revealed more differentially expressed genes, than between infant and adult blood. Functional enrichment analysis revealed an over-representation of genes involved in cell cycle and replication in thymic T cells, whereas infant blood T cells were dominated by immune related terms. Comparing adult and infant blood T cells, the former was enriched for inflammatory response, cytokine production and biological adhesion, while upregulated genes in infant blood T cells were associated with cell cycle, cell death and gene expression. CONCLUSION: This study provides valuable insight into the transcriptomes of the human primary SP T cells still residing within the thymus, and offers a unique comparison to primary blood derived T cells. Interestingly, the majority of autoimmune disease associated genes were expressed in one or more T cell subset, however ~ 11% of these were not expressed in frequently studied adult peripheral blood.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Timo/imunologia , Transcriptoma , Adulto , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular , Criança , Perfilação da Expressão Gênica/métodos , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Timo/metabolismo
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